Thyroid hormones are key regulators of basal metabolic state and oxidative metabolism. Hyperthyroidism has been reported to cause significant alterations in hemodynamics, and in cardiac and diaphragm muscle function, all of which have been linked to increased oxidative stress. However, the definite source of increased reactive oxygen species (ROS) in each of these phenotypes is still unknown. The goal of the current study was to test the hypothesis that thyroxin (T4) may produce distinct hemodynamic, cardiac, and diaphragm muscle abnormalities by differentially affecting various sources of ROS. Wild-type and T4 mice with and without 2-week treatments with allopurinol (xanthine oxidase inhibitor), apocynin (NADPH oxidase inhibitor), L-NIO (nitric oxide synthase inhibitor), or MitoTEMPO (mitochondria-targeted antioxidant) were studied. Blood pressure and echocardiography were noninvasively evaluated, followed by ex vivo assessments of isolated heart and diaphragm muscle functions. Treatment with L-NIO attenuated the T4-induced hypertension in mice. However, apocynin improved the left-ventricular (LV) dysfunction without preventing the cardiac hypertrophy in these mice. Both allopurinol and MitoTEMPO reduced the T4-induced fatigability of the diaphragm muscles. In conclusion, we show here for the first time that T4 exerts differential effects on various sources of ROS to induce distinct cardiovascular and skeletal muscle phenotypes. Additionally, we find that T4-induced LV dysfunction is independent of cardiac hypertrophy and NADPH oxidase is a key player in this process. Furthermore, we prove the significance of both xanthine oxidase and mitochondrial ROS pathways in T4-induced fatigability of diaphragm muscles. Finally, we confirm the importance of the nitric oxide pathway in T4-induced hypertension.

Force development at resting heart rate is not impacted by cardiac pathology, but kinetics are impaired and the magnitude of the impairment depends on the underlying etiology. Focusing on restoration of myocardial kinetics will likely have greater therapeutic potential than targeting force of contraction.

Studies have shown that exposure to psychological stressors leads to inflammation throughout the body. This has been widely studied using social disruption stress (SDR), a social stressor, which involves repeated social defeat in subordinate mice. Exposure to SDR increases serum cytokine levels, results in accumulation of spleen CD11b+ myeloid cells, and primes macrophages for increased cytokine and microbicidal activity. Our previous studies showed that intestinal microbes are necessary for SDR-enhancement of innate immunity. Here, we show that SDR increases spleen CD11b+Ly6CintermLy6G+ neutrophil and CD11b+Ly6ChiLy6G− monocyte numbers compared to control mice. Further, we found that neutrophils and monocytes from stressor-exposed mice expressed higher levels of IL-1β mRNA. To determine whether bacterial translocation may contribute to these effects, bacterial 16S rRNA was quantified using qRT-PCR with bacterial group-specific primers. Exposure to the SDR stressor specifically increased Lactobacillus RNA in the spleen, which localized in spleen monocytes. The increased spleen levels of Lactobacillus 16S rRNA in SDR mice positively correlated with increased levels of IL-1β and IL-23 mRNA. Our findings indicate that during stressor exposure, Lactobacillus spp. can translocate to the spleen and prime the innate immune system for enhanced reactivity.

Non-typeable Haemophilus influenzae (NTHI) is a common commensal bacterium that resides in the human upper respiratory tract of healthy individuals. NTHI is also a known causative agent of multiple diseases including sinusitis, otitis media, as well as exacerbates disease severity of patients with cystic fibrosis and chronic obstructive pulmonary disease. We have previously shown that the Sap transporter mediates resistance to host antimicrobial peptides (AMPs) and import of the iron-containing compound heme. Here, we analyzed the contribution of the Sap structural ATPase protein, SapF, in these essential functions. In contrast to SapD, SapF was dispensable for NTHI survival when exposed to AMPs in vitro. SapF was responsible for heme utilization and recovery of depleted internal heme-iron stores. Further, a loss of SapF resulted in morphological plasticity and enhanced community development and biofilm architecture, suggesting the potential role of heme-iron availability in coordinating the complexity of NTHI biofilm architecture. SapF was required for colonization of the nasopharynx and acute infection of the middle ear, as SapF deficiency correlated with a statistically significant decrease in NTHI persistence in vivo. These data suggest that SapF is required for proper heme utilization which directly impacts NTHI survival. Thus, these studies further support a role for the Sap complex in the transport of multiple substrates and further defines substrate specificity for the two ATPase subunits. Given the multiple essential functions provided by the Sap transporter, this complex could prove to be an effective therapeutic target for the treatment of NTHI diseases.

Exposure to the social disruption (SDR) stressor, which involves repeated social defeat in subordinate mice, increases serum cytokine levels, results in accumulation of CD11b+ myeloid cells in the spleen, and primes macrophages for increased cytokine and microbicidal activity. Our previous studies showed that the intestinal microbiota are necessary for SDR-enhancement of splenic macrophage microbicidal activity. The current study determined whether SDR-induced translocation of microbes to the spleen was associated with changes in spleen myeloid cell populations and cytokine mRNA. The SDR stressor increased the numbers of spleen neutrophils and inflammatory monocytes compared to control mice, while having no effect on numbers of non-classical monocytes and red pulp macrophages. Using an RNA flow cytometry assay, we found that neutrophils and inflammatory monocytes from SDR mice express higher levels of IL-1β mRNA compared to control mice. To detect translocated bacteria, qRT-PCR was used with bacterial group specific primers to 16S rRNA in addition to traditional bacterial cultures. SDR induced the translocation of Lactobacillus to the spleen, but translocation of other taxa, such as Bacteroides and Bifidobacterium, was not increased in stressor-exposed mice. Levels of Lactobacillus 16s rRNA in the spleen positively correlated with increased levels of IL-1β and IL-23 mRNA in stressor-exposed mice. Our results indicate that stressor exposure increases translocation of Lactobacillus from the gut to the spleen and primes the innate immune system for enhanced reactivity. These studies suggest that commensal microbes are key players in stressor-induced immunopotentiation. Funded by NIH grant R21AI107238.